Inhibitors of IMPDH enzyme

ABSTRACT

The present invention relates to compounds which inhibit IMPDH. This invention also relates to pharmaceutical compositions comprising these compounds. The compounds and pharmaceutical compositions of this invention are particularly well suited for inhibiting IMPDH enzyme activity and consequently, may be advantageously used as therapeutic agents for IMPDH-mediated processes. This invention also relates to methods for inhibiting the activity of IMPDH using the compounds of this invention and related compounds.

CROSS REFERENCE TO RELATED APPLICATIONS

This continuation application claims the benefit, under 35 U.S.C. §120,of U.S. patent application Ser. No. 10/997,810 filed Nov. 24, 2004 nowU.S. Pat. No. 7,432,290, which is a divisional of U.S. patentapplication Ser. No. 10/287,405 filed Nov. 4, 2002, now U.S. Pat. No.7,087,642 issued Aug. 8, 2006, which is a divisional of U.S. patentapplication Ser. No. 09/955,626 filed Sep. 19, 2001, now U.S. Pat. No.6,498,178 issued on Dec. 24, 2002, which is a continuation ofInternational Application No. PCT/US2000/07129 filed Mar. 17, 2000,which claims the priority of U.S. Application Nos. 60/125,507 filed Mar.19, 1999 and 60/174,882 filed Jan. 7, 2000. Each of the aboveapplications being incorporated herein by reference.

TECHNICAL FIELD OF THE INVENTION

The present invention relates to compounds which inhibit IMPDH. Thisinvention also relates to pharmaceutical compositions comprising thesecompounds. The compounds and pharmaceutical compositions of thisinvention are particularly well suited for inhibiting IMPDH enzymeactivity and consequently, may be advantageously used as therapeuticagents for IMPDH-mediated processes. This invention also relates tomethods for inhibiting the activity of IMPDH using the compounds of thisinvention and related compounds.

BACKGROUND OF THE INVENTION

The synthesis of nucleotides in organisms is required for the cells inthose organisms to divide and replicate. Nucleotide synthesis in mammalsmay be achieved through one of two pathways: the de novo synthesispathway or the salvage pathway. Different cell types use these pathwaysto a different extent.

Inosine-5′-monophosphate dehydrogenase (IMPDH; EC 1.1.1.205) is anenzyme involved in the de novo synthesis of guanine nucleotides. IMPDHcatalyzes the NAD-dependent oxidation of inosine-5′-monophosphate (IMP)to xanthosine-5′-monophosphate (XMP) [Jackson R. C. et. al., Nature,256, pp. 331-333, (1975]).

IMPDH is ubiquitous in eukaryotes, bacteria and protozoa [Y. Natsumeda &S. F. Carr, Ann. N.Y. Acad., 696, pp. 88-93 (1993)]. The prokaryoticforms share 30-40% sequence identity with the human enzyme. Two isoformsof human IMPDH, designated type I and type II, have been identified andsequenced [F. R. Collart and E. Huberman, J. Biol. Chem., 263, pp.15769-15772, (1988); Y. Natsumeda et. al., J. Biol. Chem., 265, pp.5292-5295, (1990)]. Each is 514 amino acids, and they share 84% sequenceidentity. Both IMPDH type I and type II form active tetramers insolution, with subunit molecular weights of 56 kDa [Y. Yamada et. al.,Biochemistry, 27, pp. 2737-2745 (1988)].

The de novo synthesis of guanosine nucleotides, and thus the activity ofIMPDH, is particularly important in B and T-lymphocytes. These cellsdepend on the de novo, rather than salvage pathway to generatesufficient levels of nucleotides necessary to initiate a proliferativeresponse to mitogen or antigen [A. C. Allison et. al., Lancet II, 1179,(1975) and A. C. Allison et. al., Ciba Found. Symp., 48, 207, (1977)].Thus, IMPDH is an attractive target for selectively inhibiting theimmune system without also inhibiting the proliferation of other cells.

Immunosuppression has been achieved by inhibiting a variety of enzymesincluding for example, the phosphatase calcineurin (inhibited bycyclosporin and FK-506); dihydroorotase dehydrogenase, an enzymeinvolved in the biosynthesis of pyrimidines (inhibited by leflunomideand brequinar); the kinase FRAP (inhibited by rapamycin); and the heatshock protein hsp70 (inhibited by deoxyspergualin). [See B. D. Kahan,Immunological Reviews, 136, pp. 29-49 (1993); R. E. Morris, The Journalof Heart and Lung Transplantation, 12(6), pp. S275-S286 (1993)].

Inhibitors of IMPDH are also known. U.S. Pat. Nos. 5,380,879 and5,444,072 and PCT publications WO 94/01105 and WO 94/12184 describemycophenolic acid (MPA) and some of its derivatives as potent,uncompetitive, reversible inhibitors of human IMPDH type I (K_(i)=33 nM)and type II (K_(i)=9 nM). MPA has been demonstrated to block theresponse of B and T-cells to mitogen or antigen [A. C. Allison et. al.,Ann. N.Y. Acad. Sci., 696, 63, (1993).

Immunosuppressants, such as MPA, are useful drugs in the treatment oftransplant rejection and autoimmune diseases. [R. E. Morris, KidneyIntl., 49, Suppl. 53, S-26, (1996)]. However, MPA is characterized byundesirable pharmacological properties, such as gastrointestinaltoxicity. [L. M. Shaw, et. al., Therapeutic Drug Monitoring, 17, pp.690-699, (1995)].

Nucleoside analogs such as tiazofurin, ribavirin and mizoribine alsoinhibit IMPDH [L. Hedstrom, et. al. Biochemistry, 29, pp. 849-854(1990)]. These compounds, however, suffer from lack of specificity toIMPDH.

Mycophenolate mofetil, a prodrug which quickly liberates free MPA invivo, was recently approved to prevent acute renal allograft rejectionfollowing kidney transplantation. [L. M. Shaw, et. al., Therapeutic DrugMonitoring, 17, pp. 690-699, (1995); H. W. Sollinger, Transplantation,60, pp. 225-232 (1995)]. Several clinical observations, however, limitthe therapeutic potential of this drug. [L. M. Shaw, et. al.,Therapeutic Drug Monitoring, 17, pp. 690-699, (1995)]. MPA is rapidlymetabolized to the inactive glucuronide in vivo. [A. C. Allison and E.M. Eugui, Immunological Reviews, 136, pp. 5-28 (1993)]. The glucuronidethen undergoes enterohepatic recycling causing accumulation of MPA inthe gastrointestinal tract where it cannot exert its IMPDH inhibitoryactivity on the immune system. This effectively lowers the drug's invivo potency, while increasing its undesirable gastrointestinal sideeffects.

More recently, IMPDH inhibitors of different classes have been describedin PCT publications WO 97/40028 and WO 98/40381.

It is also known that IMPDH plays a role in other metabolic events.Increased IMPDH activity has been observed in rapidly proliferatinghuman leukemic cell lines and other tumor cell lines, indicating IMPDHas a target for anti-cancer as well as immunosuppressive chemotherapy[M. Nagai et. al., Cancer Res., 51, pp. 3886-3890, (1991)]. IMPDH hasalso been shown to play a role in the proliferation of smooth musclecells, indicating that inhibitors of IMPDH, such as MPA or rapamycin,may be useful in preventing restenosis or other hyperproliferativevascular diseases [C. R. Gregory et al., Transplantation, 59, pp. 655-61(1995); PCT publication WO 94/12184; and PCT publication WO 94/01105].

Additionally, IMPDH has been shown to play a role in viral replicationin some virus-infected cell lines. [S. F. Carr, J. Biol. Chem., 268, pp.27286-27290 (1993)]. Analogous to lymphocytes and lymphocytic and tumorcell lines, the implication is that the de novo, rather than thesalvage, pathway is critical in the process of viral replication.

Thus, there remains a need for potent IMPDH inhibitors with improvedpharmacological properties. Such inhibitors would have therapeuticpotential as immunosuppressants, anti-cancer agents, anti-vascularhyperproliferative agents, anti-inflammatory agents, antifungal agents,antipsoriatic and anti-viral agents.

SUMMARY OF THE INVENTION

The present invention provides compounds, and pharmaceuticallyacceptable derivatives thereof, that are useful as inhibitors of IMPDH.The compounds of this invention can be used alone or in combination withother therapeutic or prophylactic agents, such as anti-virals,anti-inflammatory agents, antibiotics, and immunosuppressants for thetreatment or prophylaxis of transplant rejection and autoimmune disease.

Additionally, these compounds are useful, alone or in combination withother agents, as therapeutic and prophylactic agents for antiviral,anti-tumor, anti-cancer, anti-inflammatory agents, antifungal agents,antipsoriatic immunosuppressive chemotherapy and restenosis therapyregimens.

The invention also provides pharmaceutical compositions comprising thecompounds of this invention, as well as multi-component compositionscomprising additional IMPDH compounds together with animmunosuppressant. The invention also provides methods of using thecompounds of this invention, as well as other related compounds, for theinhibition of IMPDH.

DETAILED DESCRIPTION OF THE INVENTION

In order that the invention herein described may be more fullyunderstood, the following detailed description is set forth. In thedescription, the following abbreviations are used:

Designation Reagent or Fragment Ac acetyl Me methyl Et ethyl Bn benzylCDI carbonyldiimidazole DBU 1,8-diazabicyclo[5.4.0]undec-7-ene DIEAdiisopropylethylamine DMAP dimethylaminopyridine DMF dimethylformamideDMSO dimethylsulfoxide DPPA diphenyl phosphoryl acid EDC1-(3-dimethylaminopropyl)-3- ethylcarbodiimide hydrochloride EtOAc ethylacetate IPA isopropyl alcohol MeCN acetonitrile THF tetrahydrofuran TEAtriethylamine t-bu tert-butyl BOC butyloxycarbonyl

The following terms are employed herein:

Unless expressly stated to the contrary, the terms “—SO₂—” and “—S(O)₂—”as used herein refer to a sulfone or sulfone derivative (i.e., bothappended groups linked to the S), and not a sulfinate ester.

The terms “halo” or “halogen” refer to a radical of fluorine, chlorine,bromine or iodine.

The term “immunosuppressant” refers to a compound or drug whichpossesses immune response inhibitory activity. Examples of such agentsinclude cyclosporin A, FK506, rapamycin, leflunomide, deoxyspergualin,prednisone, azathioprine, mycophenolate mofetil, OKT3, ATAG, interferonand mizoribine.

The term “interferon” refers to all forms of interferons, including butnot limited to alpha, beta and gamma forms.

IMPDH-mediated disease refers to any disease state in which the IMPDHenzyme plays a regulatory role in the metabolic pathway of that disease.Examples of IMPDH-mediated disease include transplant rejection andautoimmune diseases, such as rheumatoid arthritis, multiple sclerosis,juvenile diabetes, asthma, and inflammatory bowel disease, as well asinflammatory diseases, cancer, viral replication diseases and vasculardiseases.

For example, the compounds, compositions and methods of using them ofthis invention may be used in the treatment of transplant rejection(e.g., kidney, liver, heart, lung, pancreas (islet cells), bone marrow,cornea, small bowel and skin allografts and heart valve xenografts),rheumatoid arthritis, multiple sclerosis, juvenile diabetes, asthma,inflammatory bowel disease (Crohn's disease, ulcerative colitis), lupus,diabetes mellitus, myasthenia gravis, psoriasis, dermatitis, eczema,seborrhea, pulmonary inflammation, eye uveitis, hepatitis, Grave'sdisease, Hashimoto's thyroiditis, Behcet's or Sjorgen's syndrome (dryeyes/mouth), pernicious or immunohaemolytic anemia, idiopathic adrenalinsufficiency, polyglandular autoimmune syndrome, andglomerulonephritis, scleroderma, lichen planus, viteligo (depigmentationof the skin), autoimmune thyroiditis, and alveolitis, inflammatorydiseases such as osteoarthritis, acute pancreatitis, chronicpancreatitis, asthma and adult respiratory distress syndrome, as well asin the treatment of cancer and tumors, such as solid tumors, lymphomasand leukemia, vascular diseases, such as restenosis, stenosis andatherosclerosis, and DNA and RNA viral replication diseases, such asretroviral diseases, and herpes.

Additionally, IMPDH enzymes are also known to be present in bacteria andthus may regulate bacterial growth. As such, the IMPDH-inhibitorcompounds, compositions and methods described herein may be useful intreatment or prevention of bacterial infection, alone or in combinationwith other antibiotic agents.

The term “treating” as used herein refers to the alleviation of symptomsof a particular disorder in a patient or the improvement of anascertainable measurement associated with a particular disorder. As usedherein, the term “patient” refers to a mammal, including a human.

The terms “HBV”, “HCV” and “HGV” refer to hepatitis-B virus, hepatitis-Cvirus and hepatitis-G virus, respectively.

According to one embodiment, the invention provides compounds of formulaA:

wherein:

-   -   each of R₁ and R₂ is independently selected from hydrogen; —CF₃;        —(C₁-C₆)-straight or branched alkyl; —(C₂-C₆)-straight or        branched alkenyl or alkynyl; —(C₁-C₆)-straight or branched        alkyl-R₇; —[(C₂-C₆)-straight or branched alkenyl or alkynyl]-R₇        or —R₇; and wherein at least one of R₁ or R₂ is (C₁-C₆)-straight        or branched alkyl-R₇; —[(C₂-C₆)-straight or branched alkenyl or        alkynyl]-R₇ or —R₇    -   wherein up to 4 hydrogen atoms in any of said alkyl, alkenyl or        alkynyl are optionally and independently replaced by R₃; or    -   wherein R₁ and R₂ are alternatively taken together to form        tetrahydrofuranyl, wherein when R₉ is hydrogen, (R)-methyl,        (R)-ethyl or (R)-hydroxymethyl, one hydrogen atom in said        tetrahydrofuran is replaced by —OR₆ or —R₇, and wherein when R₉        is (S)-methyl, (S)-ethyl or (S)-hydroxymethyl, one hydrogen atom        in said tetrahydrofuran is optionally replaced by —OR₆ or —R₇;    -   wherein when R₉ is hydrogen, (R)-methyl, (R)-ethyl or        (R)-hydroxymethyl and each of R₁ and R₂ are independently        hydrogen, unsubstituted —(C₁-C₆)-straight or branched alkyl, or        unsubstituted —(C₂-C₆)-straight or branched alkenyl or alkynyl,        then the portion of the compound represented by —CH(R₁)R₂ is a        C₅-C₁₂ straight or branched alkyl, alkenyl or alkynyl;    -   each R₃ is independently selected from halo, CN, —OR₄, or        —N(R₅)₂;    -   R₄ is selected from hydrogen, —(C₁-C₆)-straight or branched        alkyl, —(C₂-C₆)-straight or branched alkenyl or alkynyl,        —[(C₁-C₆)-straight or branched alkyl]-R₇, —[(C₂-C₆)-straight or        branched alkenyl or alkynyl]-R₇, —C(O)—[(C₁-C₆)-straight or        branched alkyl], —C(O)—[(C₂-C₆)-straight or branched alkenyl or        alkynyl], —C(O)—[(C₁-C₆)-straight or branched alkyl]-N(R₈)₂,        —C(O)—[(C₂-C₆)-straight or branched alkenyl or alkynyl]-N(R₈)₂,        —P(O) (OR₈)₂, —P(O) (OR₈)(R₈), —C(O)—R₇, —[(C₁-C₆)-straight or        branched alkyl]-CN, —S(O)₂N(R₅)₂ or —[(C₂-C₆)-straight or        branched alkenyl or alkynyl]-CN;    -   each R₅ is independently selected from hydrogen,        —(C₁-C₆)-straight or branched alkyl, —(C₂-C₆)-straight or        branched alkenyl or alkynyl, —[(C₁-C₆)-straight or branched        alkyl]-R₇, —[(C₂-C₆)-straight or branched alkenyl or        alkynyl]-R₇, —[(C₁-C₆)-straight alkyl]-CN, —[(C₂-C₆)-straight or        branched alkenyl or alkynyl]-CN, —[(C₁-C₆)-straight or branched        alkyl]-OR₄, —[(C₂-C₆)-straight or branched alkenyl or        alkynyl]-OR₄, —C(O)— (C₁-C₆)-straight or branched alkyl,        —C(O)—[(C₂-C₆)-straight or branched alkenyl or alkynyl],        —C(O)—R₇, —C(O)O—R₇, —C(O)O—(C₁-C₆)-straight or branched alkyl,        —C(O)O—[(C₂-C₆)-straight or branched alkenyl or alkynyl],        —S(O)₂—(C₁-C₆)-straight or branched alkyl, or —S(O)₂—R₇; or two        R₅ moieties, when bound to the same nitrogen atom, are taken        together with said nitrogen atom to form a 3 to 7-membered        heterocyclic ring, wherein said heterocyclic ring optionally        contains 1 to 3 additional heteroatoms independently selected        from N, O, S, S(O) or S(O)₂;    -   R₆ is selected from —C(O)—CH₃, —CH₂—C(O)—OH, —CH₂—C(O)—O-tBu,        —CH₂—CN, or —CH₂—C—CH;    -   each R₇ is a monocyclic or bicyclic ring system wherein in said        ring system:

i. each ring comprises 3 to 7 ring atoms independently selected from C,N, O or S;

ii. no more than 4 ring atoms are selected from N, O or S;

iii. any CH₂ is optionally replaced with C(O);

iv. any S is optionally replaced with S(O) or S(O)₂;

each R₈ is independently selected from hydrogen or —[C₁-C₄]-straight orbranched alkyl;

wherein in any ring system in said compound up to 3 hydrogen atoms boundto the ring atoms are optionally and independently replaced with halo,hydroxy, nitro, cyano, amino, (C₁-C₄)-straight or branched alkyl;O—(C₁-C₄)-straight or branched alkyl, (C₂-C₄)-straight or branchedalkenyl or alkynyl, or O—(C₂-C₄)-straight or branched alkenyl oralkynyl; and

-   -   wherein any ring system is optionally benzofused;    -   R₉ is selected from hydrogen, (R)-methyl, (S)-methyl, (R)-ethyl,        (S)-ethyl, (R)-hydroxymethyl or (S)-hydroxymethyl;    -   R₁₀ is selected from —C═N or 5-oxazolyl; and    -   R₁₁ is selected from halo, —O—(C₁-C₃) straight alkyl, or        —O—(C₂-C₃) straight alkenyl or alkynyl.    -   Also within the scope of formula (A) are prodrugs, which are        formed by esterifying either or both of R₁ or R₂. Examples of        such prodrugs are compounds 143 to 156 in Table 1, set forth        below.    -   The term “monocyclic ring system”, as used herein, includes        saturated, partially unsaturated and fully unsaturated ring        structures. The term “bicyclic ring system”, as used herein,        includes systems wherein each ring is independently saturated,        partially unsaturated and fully unsaturated. Examples of        monocyclic and bicyclic ring systems useful in the compounds of        this invention include, but are not limited to, cyclopentane,        cyclopentene, indane, indene, cyclohexane, cyclohexene,        cyclohexadiene, benzene, tetrahydronaphthalene,        decahydronaphthalene, naphthalene, pyridine, piperidine,        pyridazine, pyrimidine, pyrazine, 1,2,3-triazine,        1,2,4-triazine, 1,3,5-triazine, 1,2,3,4-tetrazine,        1,2,4,5-tetrazine, 1,2,3,4-tetrahydroquinoline, quinoline,        1,2,3,4-tetrahydroisoquinoline, isoquinoline, cinnoline,        phthalazine, quinazoline, quinoxaline, 1,5-naphthyridine,        1,6-naphthyridine, 1,7-naphthyridine, 1,8-naphthyridine,        2,6-naphthyridine, 2,7-naphthyridine, pteridine, acridine,        phenazine, 1,10-phenatroline, dibenzopyrans, 1-benzopyrans,        phenothiazine, phenoxazine, thianthrene, dibenzo-p-dioxin,        phenoxathiin, phenoxthionine, morpholine, thiomorpholine,        tetrahydropyran, pyran, benzopyran, 1,4-dioxane, 1,3-dioxane,        dihydropyridine, dihydropyran, 1-pyridine, quinuclidine,        triazolopyridine, β-carboline, indolizine, quinolizidine,        tetrahydronaphthyridine, diazaphenanthrenes, thiopyran,        tetrahydrothiopyran, benzodioxane, furan, benzofuran,        tetrahydrofuran, pyrrole, indole, thiophene, benzothiophene,        carbazole, pyrrolidine, pyrazole, isoxazole, isothiazole,        imidazole, oxazole, thiazole, 1,2,3-triazole, 1,2,4-triazole,        1,2,3-oxadiazole, 1,2,4-oxadiazole, 1,3,4 oxadiazole,        1,2,5-oxadiazole, 1,2,3-thiadiazole, 1,2,4-thiadiazole,        1,3,4-thiadiazole, 1,2,5 thiadiazole, tetrazole, benzothiazole,        benzoxazole, benzotriazole, benzimidazole, benzopyrazole,        benzisothiazole, benzisoxazole and purine.    -   Additional monocyclic and bicyclic structures falling within the        above description may be found in A. R. Katritzky, and C. W.        Rees, eds. “Comprehensive Heterocyclic Chemistry: Structure,        Reactions, Synthesis and Use of Heterocyclic Compounds, Vol.        1-8,” Pergamon Press, NY (1984), the disclosure of which is        herein incorporated by reference.    -   It should be understood that heterocycles may be attached to the        rest of the compound by any atom of the heterocycle which        results in the creation of a stable structure.

The term “ring atom”, as used herein, refers to a backbone atom thatmakes up the ring. Such ring atoms are selected from C, N, O or S andare bound to 2 or 3 other such ring atoms (3 in the case of certain ringatoms in a bicyclic ring system). The term “ring atom” does not includehydrogen.

The terms “—[(C₁-C₆)-straight or branched alkyl]-X” and“—[(C₂-C₆)-straight or branched alkenyl or alkynyl]-X”, wherein X isanything indicated as being bound to the alkyl, alkenyl or alkynyl,denotes that one or more X groups may be attached to the alkyl, alkenylor alkynyl chain at any termini.

According to one preferred embodiment, the compound has the formula (I):

wherein R₁ and R₂ are as defined above, or formula (IA):

wherein

R₉ is selected from (R)-methyl, (S)-methyl, (R)-ethyl, (S)-ethyl,(R)-hydroxymethyl or (S)-hydroxymethyl; and

R₁, R₂, R₁₀ and R₁₁ are as defined above.

According to a more preferred embodiment of formula IA, R₉ is selectedfrom (S)-methyl, (S)-ethyl, or (S)-hydroxymethyl methyl. Mostpreferably, R₉ is (S)-methyl. Compounds wherein R₉ is selected from(S)-methyl, (S)-ethyl, or (S)-hydroxymethyl methyl and wherein theportion of the compound represented by CH(R₁)R₂ is a C₁-C₄ straight orbranched alkyl, or a C₂-C₄ straight or branched alkenyl or alkynyl fallwithin the genus of compounds described in WO 97/40028. However,applicants have discovered that the presence of an (S) oriented moietyat R₉ imparts surprising and unexpectedly increased IMPDH inhibitoryactivity.

According to another preferred embodiment of formula IA, R₁₁ is selectedfrom O-methyl, O-ethyl or O-isopropyl.

According to a more preferred embodiment of formulae (I) and (IA), atleast one of R₁ or R₂ is selected from hydrogen, methyl, ethyl,n-propyl, isopropyl, n-butyl, t-butyl, n-pentyl, phenyl, pyridyl,—CH₂OCH₃, —CH₂CN, —CH₂OCH₂CH₂CN, —CH₂C(CH₃)₂CH₂CH₂CN,—CH₂C(CH₂CH₃)₂CH₂CH₂CN, —CH₂CH₂CN, —CH₂N(CH₂CH₂CN)₂, —CH₂N(CH₃)CH₂CH₂CN,—CH(NH₂)CH₂CN, —CH₂Cl, —CH₂OH, —CH₂CH₂OH, —CH₂CH₂OH, —CH₂CH₂CH₂CH₂OH,—CH₂CH₂OC(O)CH₃, —CH₂CH₂OC(O)CH₂NH₂, —CH₂CH₂NHCH₃, —CH₂CH₂N(CH₃)₂,—CH₂CH₂N(CH₂CH₃)₂, —CH₂N(CH₂CH₃)₂, —CH₂CH₂CH₂N(CH₃)₂,—CH₂CH₂CH₂N⁺(CH₃)₃, —CH₂OCH₂CH(CH₃)₂, —CH₂CH₂N(CH₃)C(O)OC(CH₃)₃,—CH₂N(CH₂CH₂CN)CH₂CH(CH₃)₂, —CH(CH₂CN)N(CH₃)₂,—CH₂CH(CH₂CN)NHC(O)OC(CH₃)₃,

wherein n is 0 or 1.

According to an even more preferred embodiment of formula IA, one of R₁or R₂ is selected from hydrogen, ethyl or phenyl; and the other of R₁ orR₂ is selected from —CH₂OH, —CH₂CN, —CH₂CH₂CN or CH₂N(CH₂CH₃)₂; or R₁and R₂ are taken together to form a 3-tetrahydrofuranyl moiety.

According to an alternate preferred embodiment of formula I, R₁ and R₂are taken together to form a 3-tetrahydrofuranyl moiety that issubstituted by —OR₆.

According to another preferred embodiment, the compound of formula A isselected from any of those set forth in Table 1, below.

TABLE 1 Compounds 1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

32

33

34

35

36

37

38

39

40

41

42

43

44

45

46

47

48

49

50

51

52

53

54

55

56

57

58

59

60

61

62

63

64

65

66

67

68

69

70

71

72

73

74

75

76

77

78

79

80

81

82

83

84

85

86

87

88

89

90

91

92

93

94

95

96

97

98

99

100

101

102

103

104

105

106

107

108

109

110

111

112

113

114

115

116

117

118

119

120

121

122

123

124

125

126

127

128

129

130

131

132

133

134

135

136

137

138

139

140

141

142

143

144

145

146

147

148

149

150

151

152

153

154

155

156

157

158

159

160

161

162

163

164

165

166

167

168

169

170

171

172

173

174

175

176

177

178

179

180

181

182

183

184

185

186

187

In the above table, certain compounds are shown as salts. It should beunderstood that the scope of the compounds set forth in any given entryin the table covers all forms of the depicted compound, not just thesalt shown.

When stereochemistry is not specifically indicated, the compounds ofthis invention may contain one or more asymmetric carbon atoms and thusmay occur as racemates and racemic mixtures, single enantiomers,diastereomeric mixtures and individual diastereomers. All such isomericforms of these compounds are expressly included in the presentinvention, unless otherwise indicated. Each stereogenic carbon may be ofthe R or S configuration.

Combinations of substituents and variables envisioned by this inventionare only those that result in the formation of stable compounds. Theterm “stable”, as used herein, refers to compounds that possessstability sufficient to allow manufacture and maintenance of theintegrity for a sufficient period of time to be useful for the purposesdetailed herein (e.g., therapeutic or prophylactic administration to amammal or for use in affinity chromatography applications). Typically,such compounds are stable at a temperature of 40° C. or less, in theabsence of moisture or other chemically reactive conditions, for atleast a week.

As used herein, the compounds of this invention, are defined to includepharmaceutically acceptable derivatives or prodrugs thereof. A“pharmaceutically acceptable derivative or prodrug” means anypharmaceutically acceptable salt, ester, salt of an ester, or otherderivative of a compound of this invention which, upon administration toa recipient, is capable of providing (directly or indirectly) a compoundof this invention. Particularly favored derivatives and prodrugs arethose which increase the bioavailability of the compounds of thisinvention when such compounds are administered to a mammal (e.g., byallowing an orally administered compound to be more readily absorbedinto the blood) or which enhance delivery of the parent compound to abiological compartment (e.g., the brain or lymphatic system) relative tothe parent species. Preferred prodrugs include derivatives where a groupwhich enhances aqueous solubility or active transport through the gutmembrane is appended to the structure of the compounds of thisinvention.

Pharmaceutically acceptable salts of the compounds of this inventioninclude those derived from pharmaceutically acceptable inorganic andorganic acids and bases. Examples of suitable acid salts includeacetate, adipate, alginate, aspartate, benzoate, benzene sulfonate,bisulfate, butyrate, citrate, camphorate, camphor sulfonate,cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate,fumarate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate,hexanoate, hydrochloride, hydrobromide, hydroiodide,2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate,2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectinate,persulfate, 3-phenyl-propionate, picrate, pivalate, propionate,succinate, tartrate, thiocyanate, tosylate and undecanoate. Base saltsinclude ammonium salts, alkali metal salts, such as sodium and potassiumsalts, alkaline earth metal salts, such as calcium and magnesium salts,salts with organic bases, such as dicyclohexylamine salts,N-methyl-D-glucamine, and salts with amino acids such as arginine,lysine, and so forth.

Also, the basic nitrogen-containing groups can be quaternized with suchagents as lower alkyl halides, such as methyl, ethyl, propyl, and butylchloride, bromides and iodides; dialkyl sulfates, such as dimethyl,diethyl, dibutyl and diamyl sulfates, long chain halides such as decyl,lauryl, myristyl and stearyl chlorides, bromides and iodides, aralkylhalides, such as benzyl and phenethyl bromides and others. Water oroil-soluble or dispersible products are thereby obtained.

The compounds of this invention may be synthesized using conventionaltechniques. Advantageously, these compounds are conveniently synthesizedfrom readily available starting materials. More specifically, thecompounds of this invention may be synthesized by the schemes set forthin Examples 1 and 2 with modifications that will be readily apparent tothose of skill in the art.

The compounds of this invention may be modified by appending appropriatefunctionalities to enhance selective biological properties. Suchmodifications are known in the art and include those which increasebiological penetration into a given biological compartment (e.g., blood,lymphatic system, central nervous system), increase oral availability,increase solubility to allow administration by injection, altermetabolism and alter rate of excretion.

The novel compounds of the present invention are excellent ligands forIMPDH. Accordingly, these compounds are capable of targeting andinhibiting IMPDH enzyme. Inhibition can be measured by various methods,including, for example, IMP dehydrogenase HPLC assays (measuringenzymatic production of XMP and NADH from IMP and NAD) and IMPdehydrogenase spectrophotometric assays (measuring enzymatic productionof NADH from NAD). [See C. Montero et al., Clinica Chimica Acta, 238,pp. 169-178 (1995)].

Compositions of this invention comprise a compound of this invention ora salt thereof; an additional agent selected from an immunosuppressant,an anti-cancer agent, an anti-viral agent, anti-inflammatory agent,antifungal agent, antibiotic, or an anti-vascular hyperproliferationcompound; and any pharmaceutically acceptable carrier, adjuvant orvehicle. Alternate compositions of this invention comprise a compound ofthis invention or a salt thereof; and a pharmaceutically acceptablecarrier, adjuvant or vehicle. Such composition may optionally comprisean additional agent selected from an immunosuppressant, an anti-canceragent, an anti-viral agent, anti-inflammatory agent, antifungal agent,antibiotic, or an anti-vascular hyperproliferation compound. Preferably,the compositions of this invention are pharmaceutical compositions.

The term “pharmaceutically acceptable carrier or adjuvant” refers to acarrier or adjuvant that may be administered to a patient, together witha compound of this invention, and which does not destroy thepharmacological activity thereof and is nontoxic when administered indoses sufficient to deliver a therapeutic amount of the compound.

Pharmaceutically acceptable carriers, adjuvants and vehicles that may beused in the pharmaceutical compositions of this invention include, butare not limited to, ion exchangers, alumina, aluminum stearate,lecithin, self-emulsifying drug delivery systems (SEDDS) such asdα-tocopherol polyethyleneglycol 1000 succinate, surfactants used inpharmaceutical dosage forms such as Tweens or other similar polymericdelivery matrices, serum proteins, such as human serum albumin, buffersubstances such as phosphates, glycine, sorbic acid, potassium sorbate,partial glyceride mixtures of saturated vegetable fatty acids, water,salts or electrolytes, such as protamine sulfate, disodium hydrogenphosphate, potassium hydrogen phosphate, sodium chloride, zinc salts,colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone,cellulose-based substances, polyethylene glycol, sodiumcarboxymethylcellulose, polyacrylates, waxes,polyethylene-polyoxypropylene-block polymers, polyethylene glycol andwool fat. Cyclodextrins such as α-, β-, and γ-cyclodextrin, orchemically modified derivatives such as hydroxyalkylcyclodextrins,including 2- and 3-hydroxypropyl-β-cyclodextrins, or other solubilizedderivatives may also be advantageously used to enhance delivery ofcompounds of this invention.

The pharmaceutical compositions of this invention may be administeredorally, parenterally, by inhalation spray, topically, rectally, nasally,buccally, vaginally or via an implanted reservoir. We prefer oraladministration or administration by injection. The pharmaceuticalcompositions of this invention may contain any conventional non-toxicpharmaceutically-acceptable carriers, adjuvants or vehicles. In somecases, the pH of the formulation may be adjusted with pharmaceuticallyacceptable acids, bases or buffers to enhance the stability of theformulated compound or its delivery form. The term parenteral as usedherein includes subcutaneous, intracutaneous, intravenous,intramuscular, intra-articular, intraarterial, intrasynovial,intrasternal, intrathecal, intralesional and intracranial injection orinfusion techniques.

The pharmaceutical compositions may be in the form of a sterileinjectable preparation, for example, as a sterile injectable aqueous oroleaginous suspension. This suspension may be formulated according totechniques known in the art using suitable dispersing or wetting agents(such as, for example, Tween 80) and suspending agents. The sterileinjectable preparation may also be a sterile injectable solution orsuspension in a non-toxic parenterally-acceptable diluent or solvent,for example, as a solution in 1,3-butanediol. Among the acceptablevehicles and solvents that may be employed are mannitol, water, Ringer'ssolution and isotonic sodium chloride solution. In addition, sterile,fixed oils are conventionally employed as a solvent or suspendingmedium. For this purpose, any bland fixed oil may be employed includingsynthetic mono- or diglycerides. Patty acids, such as oleic acid and itsglyceride derivatives are useful in the preparation of injectables, asare natural pharmaceutically-acceptable oils, such as olive oil orcastor oil, especially in their polyoxyethylated versions. These oilsolutions or suspensions may also contain a long-chain alcohol diluentor dispersant such as those described in Pharmacopeia Helvetica, Ph.Helv., or a similar alcohol, or carboxymethyl cellulose or similardispersing agents which are commonly used in the formulation ofpharmaceutically acceptable dosage forms such as emulsions and orsuspensions Other commonly used surfactants such as Tweens or Spansand/or other similar emulsifying agents or bioavailability enhancerswhich are commonly used in the manufacture of pharmaceuticallyacceptable solid, liquid, or other dosage forms may also be used for thepurposes of formulation.

The pharmaceutical compositions of this invention may be orallyadministered in any orally acceptable dosage form including, but notlimited to, capsules, tablets, emulsions and aqueous suspensions,dispersions and solutions. In the case of tablets for oral use, carriersthat are commonly used include lactose and corn starch. Lubricatingagents, such as magnesium stearate, are also typically added. For oraladministration in a capsule form, useful diluents include lactose anddried cornstarch. When aqueous suspensions and/or emulsions areadministered orally, the active ingredient may be suspended or dissolvedin an oily phase and combined with emulsifying and/or suspending agents.If desired, certain sweetening and/or flavoring and/or coloring agentsmay be added.

The pharmaceutical compositions of this invention may also beadministered in the form of suppositories for rectal administration.These compositions can be prepared by mixing a compound of thisinvention with a suitable non-irritating excipient which is solid atroom temperature but liquid at the rectal temperature and therefore willmelt in the rectum to release the active components. Such materialsinclude, but are not limited to, cocoa butter, beeswax and polyethyleneglycols.

Topical administration of the pharmaceutical compositions of thisinvention is especially useful when the desired treatment involves areasor organs readily accessible by topical application. For applicationtopically to the skin, the pharmaceutical composition should beformulated with a suitable ointment containing the active componentssuspended or dissolved in a carrier. Carriers for topical administrationof the compounds of this invention include, but are not limited to,mineral oil, liquid petroleum, white petroleum, propylene glycol,polyoxyethylene polyoxypropylene compound, emulsifying wax and water.Alternatively, the pharmaceutical composition can be formulated with asuitable lotion or cream containing the active compound suspended ordissolved in a carrier with suitable emulsifying agents. Suitablecarriers include, but are not limited to, mineral oil, sorbitanmonostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol,2-octyldodecanol, benzyl alcohol and water. The pharmaceuticalcompositions of this invention may also be topically applied to thelower intestinal tract by rectal suppository formulation or in asuitable enema formulation. Topically-transdermal patches are alsoincluded in this invention.

The pharmaceutical compositions of this invention may be administered bynasal aerosol or inhalation. Such compositions are prepared according totechniques well-known in the art of pharmaceutical formulation and maybe prepared as solutions in saline, employing benzyl alcohol or othersuitable preservatives, absorption promoters to enhance bioavailability,fluorocarbons, and/or other solubilizing or dispersing agents known inthe art.

Dosage levels of between about 0.01 and about 100 mg/kg body weight perday, preferably between about 0.5 and about 75 mg/kg body weight per dayof the IMPDH inhibitory compounds described herein are useful in amonotherapy and/or in combination therapy for the prevention andtreatment of IMPDH-mediated disease. Typically, the pharmaceuticalcompositions of this invention will be administered from about 1 toabout 5 times per day or alternatively, as a continuous infusion. Suchadministration can be used as a chronic or acute therapy. The amount ofactive ingredient that may be combined with the carrier materials toproduce a single dosage form will vary depending upon the host treatedand the particular mode of administration. A typical preparation willcontain from about 5% to about 95% active compound (w/w). Preferably,such preparations contain from about 20% to about 80% active compound.

When the compositions of this invention comprise a combination of anIMPDH inhibitor of this invention and one or more additional therapeuticor prophylactic agents, both the IMPDH inhibitor and the additionalagent should be present at dosage levels of between about 10 to 100%,and more preferably between about 10 to 80% of the dosage normallyadministered in a monotherapy regimen. The additional agents may beadministered separately, as part of a multiple dose regimen, from thecompounds of this invention. Alternatively, those agents may be part ofa single dosage form, mixed together with the compounds of thisinvention in a single composition.

According to one embodiment, the pharmaceutical compositions of thisinvention comprise an additional immunosuppression agent. Examples ofadditional immunosuppression agents include, but are not limited to,cyclosporin A, FK506, rapamycin, leflunomide, deoxyspergualin,prednisone, azathioprine, mycophenolate mofetil, OKT3, ATAG, interferonand mizoribine.

According to an alternate embodiment, the pharmaceutical compositions ofthis invention may additionally comprise an anti-cancer agent. Examplesof anti-cancer agents include, but are not limited to, cisplatin,actinomycin D, doxorubicin, vincristine, vinblastine, etoposide,amsacrine, mitoxantrone, tenipaside, taxol, colchicine, cyclosporin A,phenothiazines, interferon and thioxantheres.

According to another alternate embodiment, the pharmaceuticalcompositions of this invention may additionally comprise an anti-viralagent. Examples of anti-viral agents include, but are not limited to,Cytovene, Ganciclovir, trisodium phosphonoformate, Ribavirin, d4T, ddI,AZT, and acyclovir.

According to yet another alternate embodiment, the pharmaceuticalcompositions of this invention may additionally comprise ananti-vascular hyperproliferative agent. Examples of anti-vascularhyperproliferative agents include, but are not limited to, HMG Co-Areductase inhibitors such as lovastatin, thromboxane A2 synthetaseinhibitors, eicosapentaenoic acid, ciprostene, trapidil, ACE inhibitors,low molecular weight heparin, mycophenolic acid, rapamycin and5-(3′-pyridinylmethyl)benzofuran-2-carboxylate.

Upon improvement of a patient's condition, a maintenance dose of acompound, composition or combination of this invention may beadministered, if necessary. Subsequently, the dosage or frequency ofadministration, or both, may be reduced, as a function of the symptoms,to a level at which the improved condition is retained when the symptomshave been alleviated to the desired level, treatment should cease.Patients may, however, require intermittent treatment on a long-termbasis upon any recurrence of disease symptoms.

As the skilled artisan will appreciate, lower or higher doses than thoserecited above may be required. Specific dosage and treatment regimensfor any particular patient will depend upon a variety of factors,including the activity of the specific compound employed, the age, bodyweight, general health status, sex, diet, time of administration, rateof excretion, drug combination, the severity and course of the disease,the patient's disposition to the disease and the judgment of thetreating physician.

In an alternate embodiment, this invention provides methods of treatingor preventing IMPDH-mediated disease in a mammal comprising the step ofadministrating to said mammal any of the pharmaceutical compositions andcombinations described above. If the pharmaceutical composition onlycomprises the IMPDH inhibitor of this invention as the active component,such methods may additionally comprise the step of administering to saidmammal an agent selected from an anti-inflammatory agent,immunosuppressant, an anti-cancer agent, an anti-viral agent, or ananti-vascular hyperproliferation compound. Such additional agent may beadministered to the mammal prior to, concurrently with, or following theadministration of the IMPDH inhibitor composition.

In a preferred embodiment, these methods are useful in suppressing animmune response in a mammal. Such methods are useful in treating orpreventing diseases, including, transplant rejection (e.g., kidney,liver, heart, lung, pancreas (islet cells), bone marrow, cornea, smallbowel and skin allografts and heart valve xenografts), graft versus hostdisease, and autoimmune diseases, such as rheumatoid arthritis, multiplesclerosis, juvenile diabetes, asthma, inflammatory bowel disease(Crohn's disease, ulcerative colitis), lupus, diabetes, mellitusmyasthenia gravis, psoriasis, dermatitis, eczema, seborrhea, pulmonaryinflammation, eye uveitis, Grave's disease, Hashimoto's thyroiditis,Behcet's or Sjorgen's syndrome (dry eyes/mouth), pernicious orimmunohaemolytic anemia, idiopathic adrenal insufficiency, polyglandularautoimmune syndrome, glomerulonephritis, scleroderma, lichen planus,viteligo (depigmentation of the skin), autoimmune thyroiditis, andalveolitis.

These methods comprise the step of administering to the mammal acomposition comprising a compound of this invention and apharmaceutically acceptable adjuvant. In a preferred embodiment, thisparticular method comprises the additional step of administering to saidmammal a composition comprising an additional immunosuppressant and apharmaceutically acceptable adjuvant.

Alternatively, this method comprises the step of administering to saidmammal a composition comprising a compound of this invention; anadditional immunosuppressive agent and a pharmaceutically acceptableadjuvant.

In an alternate preferred embodiment, these methods are useful forinhibiting viral replication in a mammal. Such methods are useful intreating or preventing DNA and RNA viral diseases caused by infectionfor example, by orthomyxoviruses (influenza viruses types A and B),paramyxoviruses (respiratory syncytial virus (RSV), subacute sclerosingpanencephalitis (SSPE) virus) measles and parainfluenza type 3),herpesviruses (HSV-1, HSV-2, HHV-6, HHV-7, HHV-8, Epstein Barr Virus(EBV), cytomegalovirus (HCMV) and varicella zoster virus (VZV)),retroviruses (HIV-1, HIV-2, HTLV-1, HTLV-2), flavi- and pestiviruses(yellow fever virus (YFV), hepatitis C virus (HCV), dengue fever virus,bovine viral diarrhea virus (BVDV), hepatotropic viruses (hepatitis Avirus (HAV), hepatitis B virus (HBV), hepatitis D virus (HDV), hepatitisE virus (HEV), hepatitis G virus (HGV), Crimean-Congo hemorrhagic fevervirus (CCHF), bunyaviruses (Punta Toro virus, Rift Valley fever virus(RVFV), and sandfly fever Sicilian virus), Hantaan virus, Caraparuvirus), human papilloma viruses, encephalitis viruses (La Crosse virus),arena viruses (Junin and Tacaribe virus), reovirus, vesicular stomatitisvirus, rhinoviruses, enteroviruses (polio virus, coxsackie viruses,encephalomyocarditis virus (EMC)), Lassa fever virus, and togaviruses(Sindbis and Semlike forest viruses) and poxviruses (vaccinia virus),adenoviruses, rubiola, and rubella.

These methods comprise the step of administering to the mammal acomposition comprising a compound of this invention, and apharmaceutically acceptable adjuvant. In a preferred embodiment, thisparticular method comprises the additional step of administering to saidmammal a composition comprising an additional anti-viral agent and apharmaceutically acceptable adjuvant.

Alternatively, this method comprises the step of administering to saidmammal a composition comprising a compound of this invention; anadditional anti-viral agent and a pharmaceutically acceptable adjuvant.

In another alternate preferred embodiment, these methods are useful forinhibiting vascular cellular hyperproliferation in a mammal. Suchmethods are useful in treating or preventing diseases, including,restenosis, stenosis, arteriosclerosis and other hyperproliferativevascular disease.

These methods comprise the step of administering to the mammal acomposition comprising a compound of this invention, and apharmaceutically acceptable adjuvant. In a preferred embodiment, thisparticular method comprises the additional step of administering to saidmammal a composition comprising an additional anti-vascularhyperproliferative agent and a pharmaceutically acceptable adjuvant.

Alternatively, this method comprises the step of administering to saidmammal a composition comprising a compound of this invention; anadditional anti-vascular hyperproliferative agent and a pharmaceuticallyacceptable adjuvant.

In another alternate preferred embodiment, these methods are useful forinhibiting tumors and cancer in a mammal. Such methods are useful intreating or preventing diseases, including, tumors and malignancies,such as lymphoma, leukemia and other forms of cancer.

These methods comprise the step of administering to the mammal acomposition comprising a compound of this invention, and apharmaceutically acceptable adjuvant. In a preferred embodiment, thisparticular method comprises the additional step of administering to saidmammal a composition comprising an additional anti-tumor or anti-canceragent and a pharmaceutically acceptable adjuvant.

Alternatively, this method comprises the step of administering to saidmammal a composition comprising a compound of this invention; anadditional anti-tumor or anti-cancer agent and a pharmaceuticallyacceptable adjuvant.

In another alternate preferred embodiment, these methods are useful forinhibiting inflammation and inflammatory diseases in a mammal. Suchmethods are useful in treating or preventing diseases, including,osteoarthritis, acute pancreatitis, chronic pancreatitis, asthma andadult respiratory distress syndrome.

These methods comprise the step of administering to the mammal acomposition comprising a compound of this invention, and apharmaceutically acceptable adjuvant. In a preferred embodiment, thisparticular method comprises the additional step of administering to saidmammal a composition comprising an anti-inflammatory agent and apharmaceutically acceptable adjuvant.

In order that this invention be more fully understood, the followingexamples are set forth. These examples are for the purpose ofillustration only and are not to be construed as limiting the scope ofthe invention in any way.

EXAMPLE 1 Synthesis of Compound 41 A. Synthesis of C4

To a solution of glacial acetic acid (46 mL), acetic anhydride (46 mL,485 mmole) and 2-methyl-5-nitroanisole (10.0 g, 60 mmole) at 0° C. wasadded conc. H₂SO₄ (6.9 mL) in a dropwise fashion. Upon completeaddition, CrO₃ (8.08 g, 80.8 mmole) was added portion-wise over 60 mins.Following an additional 15 mins of stirring at 0° C., the reactionmixture was poured over ice and the resulting precipitate was isolatedby filtration, rinsing with cold H₂O. Purification by flashchromatography, eluting with a gradient of 15-50% EtOAc in hexanes,provided 8.14 g (24%, 51% based on recovered starting material) C1 as awhite solid. The ¹H NMR was consistent with that of the desiredstructure.

A stirred suspension of C1 (81.94 g, 307 mmole) in dioxane (100 mL) wastreated with concentrated HCl (20 mL) and heated at reflux overnight.Upon cooling to ambient temperature, the product C2 precipitated as alight yellow crystalline solid in a yield of 40.65 g (73.1%). Thefiltrate was concentrated to a volume of ca. 80 mL and a second crop ofproduct crystals was driven from solution by the addition of hexanes,yielding 8.91 g (16.0%). Both batches were identical by ¹H NMR and TLCanalysis and were consistent with that of the desired material. Thetotal yield of C2 was 49.56 g (89.1%).

A solution of C2 (456 mg, 2.51 mmole), tosylmethyl isocyanide (490 mg,2.5 mmole) and K₂CO₃ (347 mg, 251 mmole) were dissolved in methanol andheated to reflux for 1.5 hours. The product mixture was thenconcentrated in vacuo, redissolved in CH₂Cl₂, washed with water andbrine, dried over Na₂SO₄ and again concentrated in vacuo. Purifiedproduct C3 was obtained through recrystallization (Et₂O/hexanes) toyield 375 mg (68%). The ¹H NMR was consistent with that of the desiredstructure.

A solution of C3 (4.214 g, 19.1 mmole) in EtOAc (150 mL) was treatedwith 10% Pd/C (1.05 g, 25 wt. % of C3) and subjected to 40 psi H₂(g)(Parr Hydrogenation Apparatus) overnight. The reaction mixture wasfiltered and concentrated in vacuo. Pure product C4 was obtained throughflash chromatography, eluting with a gradient of 30-40% EtOAc/hexanes,in a yield of 3.4 g (93%). The ¹H NMR was consistent with that of thedesired structure.

B. Synthesis of Compound I113

A solution of 3-aminobenzylamine (826 mg, 6.87 mmole) and triethylamine(2.39 mL, 17.18 mmole) was treated with di-t-butyldicarbonate (1.50 g,6.87 mmole) and the mixture was stirred at ambient temperature for 2hours. The reaction was then diluted with CH₂Cl₂, washed withNaHCO₃(aq), water and brine, dried (Na₂SO₄) and concentrated in vacuo.Pure E1 was obtained by flash chromatography, eluting with 25% EtOAc inhexanes in a yield of 200 mg (46%). The ¹H NMR was consistent with thatof the desired structure.

A solution of C4 (150 mg, 0.789 mmole) and 1,1-dicarbonylimidiazole (160mg, 0.986 mmole) were combined in THF (5 mL) and stirred for 6 hours atambient temperature. The precipitation of imidazole was noted. To thiswas then added E1 (351 mg, 1.58 mmole) and N,N-dimethylaminopyridine (97mg, 0.789 mmole) and the mixture was refluxed overnight, resulting in ahomogenous solution. Upon cooling to ambient temperature, the reactionwas diluted with EtOAc (20 mL), washed with KHSO₄(aq), water, and brine,dried (MgSO₄) and concentrated. Pure I113 was obtained through flashchromatography, eluting with a gradient of 20-30-35% acetone in hexanesin a yield of 164 mg (47%). ¹H NMR (500 MHz, d₆-DMSO) δ 8.90 (s), 8.75(s), 8.38 (s), 7.60 (d), 7.51 (s), 7.3-7.46 (m), 7.21-7.27 (t), 7.05(dd), 6.87 (d), 4.12 (d), 3.93 (s), 1.44 (s). R_(f) 0.21 (5%MeOH/CH₂Cl₂).

C. Synthesis of Compound I168

A suspension of I113 (250 mg, 5.76 mmol) in CH₂Cl₂ (1 mL) was treated ina dropwise fashion at ambient temperature with several equivalents oftrifluoroacetic acid and stirred for 90 min. The resulting solution wasstripped in vacuo and titrated with CH₂Cl₂ and methanol. Pure productI168 was isolated by filtration in a yield of 258 mg (99%). The ¹H NMRwas consistent with that of the desired product.

D. Synthesis of Compound 41

To a room temperature solution of 1-methoxy-2-propanol (75 mg, 832μmole) in THF (1.0 mL) was added solid 1,1′-carbonyl diimidazole (121mg, 749 μmole) in one portion. The resulting mixture was stirred at roomtemperature overnight, then treated sequentially with TEA (174 μL, 1.25mmole), solid compound I168 (376 mg, 832 μmole), and DMF (1.0 mL). Theresulting solution was stirred at room temperature for one day, thendiluted with ethyl acetate, washed sequentially with water and brine,dried over MgSO₄, filtered, and concentrated in vacuo. The crude productwas then purified by flash chromatography (silica gel, 97.5/1.5 CH₂Cl₂).The chromatographed product was then triturated with a 9/1 mixture ofethyl ether/ethyl acetate to give compound 45 (65 mg, 56% yield) as awhite, powdery solid.

¹H NMR (500 MHz, acetone-d6): 8.34 (s, 1H); 8.21 (s, 1H); 8.12 (s, 1H);7.67 (s, 1H); 7.65 (dd, 1H); 7.50 (d, 1H); 7.47 (d, 1H); 7.43 (s, 1H);7.25 (dd, 1H); 7.10 (dd, 1H); 6.97 (d, 1H); 6.68 (m, 1H); 4.92 (m, 1H);4.32 (d, 2H); 4.01 (s, 3H); 3.43 (dd, 1H); 3.33 (dd, 1H); 3.31 (s, 3H);1.18 (d, 3H).

Other compounds of this invention may be prepared in a similar mannersubstituting the appropriate alcohol for 1-methoxy-2-propanol [i.e.,HO—CH(R₁)(R₂)] in step C.

EXAMPLE 2 Preparation of Compound 169 A. Preparation of the Left HandSide Coupling Intermediate (R₁₀=cyano)

Copper(I) cyanide (7.2 g, 80.8 mmole) was combined with2-bromo-5-nitroanisole (I) (15 g, 64.6 mmole) in NMP (70 mL) and heatedto 150° C. overnight under an N₂ atmosphere. The mixture was treatedwith Celite, cooled to room temperature, then diluted with EtOAc and 1.0N NaOH and allowed to stir for 15 minutes. The heterogeneous mixture wasfiltered through a pad of Celite with EtOAc, the phases were separated,and the aqueous phase was washed 3 times with EtOAc. The combinedorganics were washed sequentially with 1.0 N NaOH, water, and brine,then dried over Na₂SO₄, filtered and concentrated in vacuo. The crudeproduct was dissolved in CH₂Cl₂, filtered through a short pad of silicagel to remove solids and most colored impurities, then concentrated invacuo to give II (10.41 g, 90%) as a brownish-orange solid.

¹H NMR (500 MHz, CDCl₃): 7.90 (d, 1H); 7.84 (s, 1H); 7.77 (d, 1H); 4.07(s, 3H).

To a room temperature solution of II (7.2 g, 40.4 mmoles) in EtOAc-EtOH(220-15 mL) was added 10% Pd/C (1.8 g) resulting in a heterogeneousblack mixture. The reaction was placed under 1 atmosphere (balloon) ofH₂, warmed to 50° C., and stirred overnight. Reaction was cooled to roomtemperature, the catalyst was removed via filtration, and the filtratewas concentrated in vacuo to give III (5.56 g, 93%) as a crystallinesolid.

¹H NMR (500 MHz, CDCl₃): 7.29 (d, 1H); 6.22 (d, 1H); 6.17 (s, 1H); 4.20(broad s, 2H); 3.85 (s, 3H).

To a room temperature, biphasic mixture of phenyl chloroformate (1.6 mL,12.82 mmoles) in EtOAc (20 mL) and sat. NaHCO₃ (˜1M, 16 mL) was addedIII (950 mg, 6.41 mmoles) as a solution in EtOAc (10 mL) over a 10minute period. The resulting heterogeneous mixture was stirred at roomtemperature for 30 minutes and then the phases were separated. Theorganic phase was washed with brine, dried over Na₂SO₄, filtered througha pad of silica gel with EtOAc, and concentrated in vacuo to give athick oil. The resulting oil was diluted in toluene (30 mL) and treatedwith hexanes (30 mL) resulting in a thick precipitate. This mixture wasstirred for 30 minutes, filtered, solids washed with 1:1toluene:hexanes, then hexanes alone, and dried to constant weight underhigh vacuum to give IV (1.65 g, 96%) as a white powder.

¹H NMR (500 MHz, dmso-d6); 10.76 (s, 1H); 7.69 (d, 1H); 7.44 (d, 1H);7.40 (d, 1H); 7.26 (m, 3H); 7.15 (d, 1H); 3.85 (s, 3H).

B. Preparation of the Right Hand Side Coupling Intermediate(R₂=S-methyl)

To a room temperature solution of V (200 g, 1.21 moles) in EtOH (2 L)was added NaBH₄ (50.3 g, 1.33 moles) portionwise over 30 minutes, notallowing the internal temperature to rise over 40° C. The reaction wasallowed to stir at room temperature for 4 hours. It was then quenchedwith water (˜100 mL), concentrated in vacuo, diluted with EtOAc, washedtwice with water, once with sat. NaHCO₃, dried over MgSO₄, filtered, andconcentrated in vacuo to give VI (191.7 g, 95%) as a yellowish power.

¹H NMR (500 MHz, CDCl₃): 8.21 (s, 1H); 8.09 (d, 1H); 7.70 (d, 1H); 7.49(dd, 1H); 5.01 (dd, 1H); 2.45 (s, 1H); 1.52 (d, 3H).

To a room temperature solution of VI (181 g, 1.08 moles) was added DPPA(250 mL, 1.16 moles) at a rate slow enough to keep the reactiontemperature under 45° C. Once the addition of DPPA was complete, themixture was treated with DBU (177 mL, 1.18 moles) at a rate slow enoughto keep the reaction temperature under 45° C. Upon complete addition,the reaction was warmed to 60° C. and maintained at that temperatureovernight. The resulting biphasic mixture was cooled to roomtemperature, washed sequentially with water, then 0.5 M HCl. The organicphase was dried over Na₂SO₄, filtered, and concentrated in vacuo to givea yellow-green oil that was not purified further.

¹H NMR (500 MHz, CDCl₃): 8.21 (s, 1H); 8.18 (d, 1H); 7.68 (d, 1H); 7.56(q, 1H); 4.76 (dd, 1H); 1.59 (d, 3H).

To a room temperature solution of VII (8.17 g, 42.51 mmoles) inTHF-water (80 mL-10 mL) was added Ph₃P (12.3 g, 46.76 mmoles) as asolution in THF (20 mL) over a 10 minute period. Nitrogen evolution wasimmediate and constant throughout the addition. The reaction was thenheated to 65° C. overnight, then cooled to room temperature. The crudemixture was concentrated in vacuo, diluted with EtOAc, washed withbrine, dried over Na₂SO₄, and filtered. The resulting filtrate wastreated with 1 N HCl/Et₂O at room temperature over a 10 minute periodresulting in precipitate formation. The mixture was stirred at roomtemperature for 15 minutes, then filtered. The solids were washed withEt₂O to give a yellow powder. The crude amine hydrochloride salt wassuspended in brine/EtOAc, and treated with 10 N NaOH (5 mL, 50 mmoles)at room temperature. The resulting mixture was stirred at roomtemperature until all solids were dissolved. The phases were separated,the aqueous phase was washed with EtOAc twice, the combined organicphases were washed with brine, dried over Na₂SO₄, filtered, andconcentrated in vacuo. The crude amine was diluted in MeOH (50 mL) andadded to a refluxing solution of L-(+)-tartaric acid (5.33 g, 35.33mmoles) in MeOH (450 mL). A precipitate formed immediately and was thendissolved in the MeOH mixture upon refluxing for 15 minutes. Theinternal temperature was lowered to 50° C. and maintained thereovernight. The internal temperature was then lowered to 30° C. andmaintained for another 24 hours followed by another 24 hours at roomtemperature. The resulting crystals (spikes) were filtered, washed withMeOH and Et₂O, and the mother liquor discarded. The resulting crystalswere dissolved in 200 mL of refluxing MeOH, cooled slowly as describedabove, filtered, and washed with MeOH, then Et₂O to give the first cropof VIII (2.21 g, 20%) as a white solid. The mother liquor wasconcentrated in vacuo, solids dissolved in 50 mL of refluxing MeOH,cooled as above, filtered, and washed with MeOH and Et₂O to give asecond crop of VIII (1.50 g, 13%) as a white solid. The optical puritywas determined on the corresponding phenyl carbamate of each crop tobe >97% ee.

Enantiomeric excesses were determined using a Chiralcel OD column (0.46cm×25 cm) made by Daicel Chemical Industries and purchased from ChiralTechnologies. The mobile phase employed was a 70:30 hexane:IPA mixturein an isocratic run out to 65 minutes at 0.8 ml/min flow rate using a3-4 μl injection of a 1-2 mg/ml solution of the phenyl carbamatedissolved in above mentioned hexane:IPA mixture. The desired S-methylenantiomer elutes first at ˜47.2 minutes while the undesired R-methylenantiomer comes off at ˜51.7 minutes while monitoring at 214, 254, 280nm wavelength.

All samples were run on a Hewlett Packard Series 1050 HPLC with a diodearray detector.

To a heterogeneous suspension of VIII (1.11 g, 3.51 mmoles) in EtOAc (20mL) and brine (20 mL) was added 10 N NaOH (0.77 mL, 7.72 mmoles) at roomtemperature. The resulting mixture was stirred at room temperature untilall salts had dissolved. The phases were then separated, and the aqueousphase washed with EtOAc. The combined organic phases were washed withbrine, dried over Na₂SO₄, filtered, and concentrated in vacuo. The crudenitro-benzylamine was diluted in 7M NH₃-MeOH (20 mL), 20% Pd(OH)₂—Cadded, and placed under 45 psi of H₂ for 5 hours. The resulting mixturewas filtered to remove the catalyst, concentrated in vacuo, azeotropedonce with CH₂Cl₂, then placed under high vacuum to give IX (455 mg, 95%)as a waxy white solid.

¹H NMR (500 MHz, dmso-d6): 6.91 (dd, 1H); 6.56 (s, 1H); 6.50 (d, 1H);6.38 (d, 1H); 4.90 (broad s, 2H); 3.82 (q, 1H); 3.31 (broad s, 2H); 1.18(d, 3H).

C. Preparation of Compound 169

To a room temperature solution of 3-(R)-hydroxy pentanitrile (212 mg,2.14 mmoles) was added CDI (521 mg, 3.21 mmoles) in one portion. Theresulting mixture was stirred at room temperature for 1 hour, thentreated with solid silica gel. The heterogeneous mixture was stirredvigorously for 10 minutes, filtered through a short pad of silica gelwith 4:1 EtOAc:IPA, concentrated in vacuo, azeotroped twice with MeCN,then combined with IX (350 mg, 2.57 mmoles) in MeCN (2 mL) and stirredat room temperature for 1 day. The resulting mixture was diluted withEtOAc, washed with water and then brine, dried over Na₂SO₄, filtered,concentrated, and flash chromatographed (silica gel, 1/2→1/3→0/1hexanes/EtOAc→4/1 EtOAc/IPA) to give X (472 mg, 84%) as a clear, thickoil.

¹H NMR (500 MHz, dmso-d6): 7.73 (d, 1H); 6.94 (dd, 1H); 6.51 (s, 1H);6.47 (d, 1H); 6.38 (d, 1H); 4.98 (broad S, 2H); 4.67 (m, 1H); 4.49 (m,1H); 2.82 (m, 2H); 1.62 (m, 2H); 1.27 (d, 3H); 0.89 (dd, 3H).

To a room temperature solution of X (470 mg, 1.80 mmoles) in EtOAc (5mL) was added IV (440 mg, 1.63 mmoles) and TEA (0.23 mL, 1.63 mmoles).The resulting mixture was heated to reflux and stirred at thattemperature for 6 hours. The resulting crude mixture was cooled to roomtemperature, diluted with EtOAc, washed with brine/1N HCl, followed bybrine alone, dried over Na₂SO₄, filtered, concentrated in vacuo, andflash chromatographed (silica gel, 1/1→1/2→1/3→1/4→0/1 hexanes/EtOAc→4/1EtOAc/IPA) to give 169 (740 mg, 100%) as a white, foamy solid.

¹H NMR (500 MHz, dmso-d6): 9.21 (s, 1H); 8.84 (s, 1H); 7.93 (d, 1H);7.59 (d, 1H); 7.51 (s, 1H); 7.41 (s, 1H); 7.29 (d, 1H); 7.23 (dd, 1H);7.01 (d, 1H); 6.92 (d, 1H); 4.69 (m, 1H); 4.63 (m, 1H); 3.89 (s, 3H);2.82 (m, 2H); 2.62 (m, 2H); 1.31 (d, 3H); 0.90 (t, 3H)

EXAMPLE 3 IMPDH Activity Inhibition Assay

IMP dehydrogenase activity was assayed following an adaptation of themethod first reported by Magasanik. [B. Magasanik et al., J. Biol.Chem., 226, p. 339 (1957), the disclosure of which is hereinincorporated by reference]. Enzyme activity was measuredspectrophotometrically, by monitoring the increase in absorbance at 340nm due to the formation of NADH (□0340 is 6220 M⁻¹ cm⁻¹). The reactionmixture contained 0.1 M potassium phosphate 8.0, 0.5 mM EDTA, 2 mM DTT,200 μM IMP and enzyme (IMPDH human type II) at a concentration of 15 to50 nM. This solution is incubated at 37° C. for 10 minutes. The reactionis started by adding NAD to a final concentration of 200 μM and theinitial rate is measured by following the linear increase in absorbanceat 340 nm for 10 minutes. For reading in a standard spectrophotometer(path length 1 cm) the final volume in the cuvette is 1.0 ml. The assayhas also been adapted to a 96 well microtiter plate format; in this casethe concentrations of all the reagents remain the same and the finalvolume is decreased to 200 μl.

For the analysis of inhibitors, the compound in question is dissolved inDMSO to a final concentration of 20 mM and added to the initial assaymixture for preincubation with the enzyme at a final volume of 2-5%(v/v). The reaction is started by the addition of NAD, and the initialrates measured as above. K_(i) determinations are made by measuring theinitial velocities in the presence of varying amounts of inhibitor andfitting the data using the tight-binding equations of Henderson(Henderson, P. J. F. (1972) Biochem. J. 127, 321].

These results are shown in Table 2. Category “A” indicates a K_(I) of 10nM or less, category “B” indicates a K_(I) of greater than 10 and lessthan 50 nM, category “C” indicates a K_(I) of 50 nM or greater, “ND”indicates inhibitory activity was not determined.

TABLE 2 IMPDH inhibitory activity. Cmpd KI (nM) 1 A 2 A 3 A 4 A 5 A 6 A7 A 8 A 9 A 10 A 11 A 12 A 13 A 14 A 15 A 16 A 17 A 18 A 19 A 20 A 21 A22 A 23 A 24 A 25 A 26 A 27 A 28 A 29 A 30 A 31 B 32 A 33 A 34 A 35 A 36A 37 A 38 A 39 A 40 A 41 A 42 A 43 A 44 A 45 A 46 A 47 A 48 A 49 A 50 A51 A 52 A 53 A 54 A 55 A 56 A 57 A 58 A 59 A 60 A 61 A 62 A 63 A 64 ND65 A 66 A 67 A 68 A 69 A 70 A 71 A 72 A 73 A 74 A 75 A 76 A 77 A 78 A 79A 80 A 81 B 82 C 83 B 84 B 85 C 86 B 87 A 88 A 89 A 90 A 91 A 92 A 93 B94 A 95 A 96 B 97 B 98 B 99 B 100 B 101 A 102 A 103 C 104 B 105 A 106 C107 C 108 C 109 C 110 C 111 C 112 C 113 C 114 C 115 C 116 C 117 C 118 C119 C 120 C 121 ND 122 C 123 C 124 C 125 C 126 C 127 C 128 C 129 C 130 C131 C 132 C 133 B 134 B 135 C 136 C 137 C 138 A 139 C 140 C 141 B 142 B143 ND 144 ND 145 ND 146 ND 147 ND 148 ND 149 ND 150 ND 151 ND 152 ND153 ND 154 ND 155 ND 156 ND 157 B 158 B 159 A 160 C 161 A 162 B 163 B164 B 165 C 166 C 167 C 168 B 169 A 170 A 171 C 172 C 172 C 173 C 174 C175 C 176 C 177 C 178 C 179 C 180 B 181 A 182 C 183 B 184 B 185 B 186 C187 B

Other compounds of this invention will also have IMPDH inhibitoryactivity.

EXAMPLE 4 Cellular Assays

A. Isolation of peripheral blood mononuclear cells (PBMCs): Human venousblood was drawn from normal healthy volunteers using heparin as ananti-coagulant. PBMCs were isolated from blood by centrifugation overFicoll-plaque gradient or CPT tubes (Becton-Dickinson) using standardconditions. PBMCs were harvested, washed and re-suspended in completeRPMI, counted and diluted to 1×10⁶ cells/mL.

B. PBMC and Splenocyte Proliferation Assays:

5×10⁴ cells (for human PBMC T cells) or 1×10⁵ cells (for human PBMC Bcells) were added per well of a 96-well plate. For T-cell assays,phyto-hemagglutinin (PHA) was added to a final concentration of 10-20μg/mL per well for cell. For B-cell assays, Staphylococcal protein A(SPAS) was added to a final concentration of 2 μg/mL per well.

Serial 4-fold dilutions of inhibitor stocks were made in complete RPMIand added to cells such that the final concentration of compounds rangedfrom 20 μM to 20 nM, while DMSO was maintained at a final concentrationof 0.1%. The cells were then incubated for 3 days. All samples weretested in triplicate. Tritiated thymidine (0.4 μCi/well) was added forthe last 24 hours of the assay. The cells were harvested onto Betaplatefilters and counted in a scintillation counter. Concentrations ofcompounds required to inhibit proliferation of cells by 50% (IC50values) were calculated using the SoftMax Pro™ (Molecular Devices)computer software package.

The results of these assays are shown in Table 3. Category “A” indicatesa IC₅₀ of 100 mM or less, category “B” indicates a IC₅₀ of greater than100 and less than 1000 nM, category “C” indicates a IC₅₀ of 1000 mM orgreater, “ND” indicates inhibitory activity was not determined in theindicated cellular assay.

TABLE 3 Cellular Activity T-cells B-cells Cmpd (IC50) (IC50) 1 B A 2 B B3 B B 4 C C 5 C C 6 B B 7 B B 8 B B 9 B B 10 B C 11 C B 12 B B 13 B B 14C B 15 B B 16 C C 17 C C 18 C C 19 B B 20 B B 21 B C 22 B B 23 A A 24 CC 25 B B 26 A A 27 A A 28 B B 29 A A 30 C B 31 ND ND 32 B A 33 B B 34 CB 35 B B 36 C C 37 B B 38 B B 39 B B 40 C B 41 B B 42 B B 43 B B 44 B B45 B B 46 B B 47 B B 48 B B 49 B B 50 C C 51 B B 52 C C 53 ND ND 54 NDND 55 ND ND 56 ND B 57 B B 58 B B 59 C B 60 B B 61 B B 62 B B 63 B B 64ND ND 65 B B 66 C C 67 B B 68 B B 69 B B 70 C C 71 C C 72 B B 73 C C 74C C 75 B B 76 A A 77 B B 78 B B 79 B B 80 B A 81 B B 82 B B 83 B B 84 BB 85 ND ND 86 C C 87 A A 88 B B 89 A A 90 B B 91 C C 92 B B 93 ND ND 94ND ND 95 ND ND 96 ND ND 97 ND ND 98 A A 99 B B 100 B B 101 A A 102 B B103 B A 104 A A 105 B B 106 A A 107 B B 108 B A 109 B B 110 B A 111 B B112 C C 113 C C 114 C C 115 C C 116 C C 117 C C 118 B B 119 B B 120 C C121 ND ND 122 C C 123 C C 125 B B 126 C C 127 C C 128 B B 129 B B 130 BC 131 B B 132 B B 133 ND ND 134 B B 135 ND ND 136 ND ND 137 ND ND 138 NDND 139 B B 140 ND ND 141 ND ND 142 ND ND 143 ND ND 144 ND ND 145 ND ND146 ND ND 147 ND ND 148 ND ND 149 ND ND 150 ND ND 151 ND ND 152 ND ND153 ND ND 154 ND ND 155 ND ND 156 ND ND 157 A A 158 C B 159 B B 160 NDND 161 ND ND 162 B B 163 A A 164 B B 165 ND ND 166 B B 167 ND ND 168 B B169 A A 170 B B 171 B A 172 C C 173 C B 174 C B 175 C C 176 C C 177 NDND 178 ND ND 179 C C 180 B C 181 A A 182 C B 183 B B 184 B B 185 B A 186ND ND 187 B A

EXAMPLE 5 Anti-Viral Assays

The anti-viral efficacy of compounds may be evaluated in various invitro and in vivo assays. For example, compounds may be tested in invitro viral replication assays. In vitro assays may employ whole cellsor isolated cellular components. In vivo assays include animal modelsfor viral diseases. Examples of such animal models include, but are notlimited to, rodent models for HBV or HCV infection, the Woodchuck modelfor HBV infection, and chimpanzee model for HCV infection.

While we have hereinbefore presented a number of embodiments of thisinvention, it is apparent that our basic construction can be altered toprovide other embodiments which utilize the methods of this invention.Therefore, it will be appreciated that the scope of this invention is tobe defined by the claims appended hereto rather than the specificembodiments which have been presented hereinbefore by way of example.

1. A method for inhibiting lymphoma or leukemia in a mammal, comprising the step of administering to said mammal a pharmaceutical composition comprising a compound of formula 181:

in an amount effective to inhibit IMPDH and a pharmaceutically acceptable carrier, adjuvant, or vehicle.
 2. The method according to claim 1, wherein said mammal is administered an additional anti-tumor or anti-cancer agent in a separate dosage form or as part of said pharmaceutical composition. 